Rotavirus is a genus of the family Reoviridae. This genus of viruses is widely recognized as the major cause of gastroenteritis of infants and young children in most areas of the world. In the lesser developed countries diarrheal diseases such as gastroenteritis constitute a major cause of mortality among infants and young children. For a general background on rotoviruses, see Kapikian et al., in Virology, pp. 863-906 (B. N. Fields et al., eds., 1985), the disclosure of which is incorporated herein by reference.
Immunity to rotavirus infections and illness has been poorly understood. Animal studies, however, have been conducted directed to the relative importance of systemic and local immunity. Bridger et al. (1981) Infect. Immun. 31:906-910; Lecce et al. (1982) J. Clin. Microbiol. 16:715-723; Little et al. (1982) Infect. Immun. 38:755-763. For example, it has been observed that calves develop a diarrheal illness despite the presence of serum rotavirus antibody at the time of infection. Calves which are fed colostrum-containing rotavirus antibodies immediately before and after infection with rotavirus, however, do not develop diarrhea within the normal incubation period. See, e.g., Bridger et al. (1975) Br. Vet. J. 131:528-535; Woode et al. (1975) Vet. Rec. 97:148-149. Similar results have been achieved with newborn lambs, who developed resistance when fed colostrum or serum containing rotavirus antibodies for several days during which period the lambs were challenged with rotavirus. Snodgrass et al. (1976) Arch. Virol. 52:201-205.
In studies of the effect of administering rotavirus to humans, it was found that a preexisting high titer of serum neutralizing antibodies to rotavirus correlated with resistance to diarrheal illness. Kapikian et al. (1983) Dev. Biol. Standard 53:209-218; Kapikian et al. (1983) J. Infect. Dis. 147:95-106. In infants and children, however, the presence of serum antibody to rotavirus has not been associated with resistance to infection or illness. See, e.g., Black et at.. (1982) J. Infect. Dis. 145:483-489; Gurwith et al. (1981) J, Infect. Dis. 144:218-224; McLean et al. (1981) J. Clin. Microbiol. 13:22-29.
Most current efforts in experimental rotavirus immunoprophylaxis are aimed at the development of live attenuated virus vaccines. Attenuation, however, is usually associated with a decrease in the level of viral replication in the target organ; i.e., the epithelium the small intestine. Attenuated mutants of other mucosal viruses, however, have exhibited a diminished immune response correlated with the decrease in replication. Since the protective efficacy of wild-type virus infection is marginal, it may be impossible to achieve the desired immunoprophylaxis with a mutant exhibit decreased replication. Two bovine rotaviruses, NCDV and the UK strain, have been produced in attenuated form and evaluated as vaccines in humans. Vesikari et al. (1983) Lancet 2:807-811; Vesikari et al. (1984) Lancet 1:977-981; Wyatt et al. (1984) in Conference Proceedings: Control and Eradication of Infectious Diseases in Latin America.
Another approach to the development of an attenuated rotavirus vaccine is based on the ability of rotaviruses to undergo gene reassortment during coinfection. A number of "hybrid" strains have been isolated from cultures coinfected with a wild-type animal rotavirus and a human rotavirus. Strains are selected which receive the gene coding for the outer nuclear capsid protein VP7, the remaining genes being derived from the animal rotavirus parent. See, e.g., Immunogenicity, pp. 319-327 (Chanock & Lerner, eds., 1984).
Still another approach to immunization has been the suggestion of using recombinantly produced VP7 polypeptide in a vaccine. See, e.g., Virology, p. 892 (B. N. Fields et al., eds., 1985). It has been further suggested, however, that recombinant VP7 is unlikely to produce an effective primary local intestinal immune response. Id. at 893. The VP7 gene from several strains of rotavirus has been cloned and full-length or near full-length cDNA has been attained. See, e.g., Arias et al. (1984) J. Virol. 50:657-661; Both et al. (1983) Proc. Natl. Acad. Sci. USA 80:3091-3095; Elleman et al. (1983) Nucleic Acid Res. 11:4689-4701; Flores et al. in Modern Approached to Vaccines; Molecular and Chemical Basis of Virus Virulence and Immunogenicity, pp. 159-164 (R. M. Chanock et al., eds., 1983).
It has also been suggested that synthetic peptides corresponding to major anogenic sites of VP7 may be useful in immunization. Virology, supra, p. 893. In addition, passive immunization with rotavirus antibodies has been shown to be effective in preventing rotavirus illness in animals and in infants and young children. Id.
The most abundant structural protein in rotavirus particles is the approximate 45K MW nucleocapsid or inner capsid protein coded for by gene known in the art as virus protein 6 or VP6. Although not an integral component of the outer capsid, it is an important viral antigen. It has been identified as the subgroup antigen by using several techniques including complement fixation, ELISA, immunoadherence agglutination assay, and specific monoclonal antibodies. VP6 is also described as the common rotavirus group antigen since some monoclonal antibodies against it will react with all rotaviruses, and polyclonal serum raised against a single rotavirus type can detect most other rotavirus strains. Aside from its antigenic properties, VP6 is very immunogenic and several investigators have found that polyclonal serum raised to this protein has neutralizing ability. Bastardo et al. (1981) Infect. & Immun. 34:641-647.
The gene encoding VP6 has been cloned. See, e.g., Estes et al. (1984) Nucleic Acids Res. 12:1875-1887. VP6 has also been produced by recombinant methods. Estes et al. (1987) J. Virol. 61:1488-1494.
Vaccine compositions for rotavirus disease comprised of peptides from VP7, VP6 and VP3 have also been proposed. See commonly owned patent applications: U.S. Ser. No. 903,325 (filed Sep. 3, 1986); Canadian Ser. No. 526,116 (filed Dec. 23, 1986); Australian Ser. No. 66987/86 (filed Dec. 24, 1986); Chinese Ser. No. 86108975 (filed Dec. 25, 1986); EPO Ser. No. 117 981.0 (Dec. 23, 1986); and Japanese Ser. No. 61-308945 (filed Dec. 26, 1986), the disclosures of which are incorporated by reference herein.
Several immunologic carriers are known in the art, including, but not limited to, keyhole limpet hemocyanin (KLH), bovine serum albumin (BSA), ovalbumin (OVA), beta-galactosidase (B-GAL), penicillinase, poly-DL-alanyl-poly-L-lysine, and poly-L-lysine. The coupling of the desired hapten or other epitope-bearing molecule to such carriers often requires elaborate chemical procedures. Such procedures are expensive and may have a deleterious effect on the final complex comprised of the carrier and epitope-bearing molecule. Thus, there is a need in the art for improved immunological carriers to which epitope-bearing molecules can be attached readily, but which are also at least as effective as prier art immunologic carriers.